LOCUS       sequence-263100-        4001 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Part of GoldenGate Yarrowia lipolytica toolkit for assembling an 
            expression plasmid for Y. lipolytic yeast. Inducible promoter for 
            gene expression in position 3 (P3_EYK300-4AB).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4001)
  AUTHORS   Larroude M, Park YK, Soudier P, Kubiak M, Nicaud JM, Rossignol T
  TITLE     A modular Golden Gate toolkit for Yarrowia lipolytica synthetic 
            biology.
  JOURNAL   Microb Biotechnol. 2019 May 31. doi: 10.1111/1751-7915.13427.
  PUBMED    31148366
REFERENCE   2  (bases 1 to 4001)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Microb 
            Biotechnol. 2019 May 31. doi: 10.1111/1751-7915.13427."
COMMENT     Sequence Label: GGE275
FEATURES             Location/Qualifiers
     source          1..4001
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    107..128
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(143..148,149..166,167..173)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    181..197
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     186..208
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     205..221
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     205..221
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        239..257
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     239..256
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(889..908)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(890..908)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(915..932)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(915..931)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(924..946)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     CDS             1069..1371
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="QFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDKV
                     SRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     1250..1269
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             1720..2514
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             1720..2514
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(1774..1793)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2384..2403
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     regulatory      2714..2723
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2720..3094
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     rep_origin      3232..3820
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3721..3740
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3974..3991
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 agcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc
       61 acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc
      121 tcactcatta ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa
      181 ttgtgagcgg ataacaattt cacacaggaa acagctatga ccatgattac gccaagctat
      241 ttaggtgaca ctatagaata ctcaagctat gcatcaagct tggtaccgag ctcggatcca
      301 ctagtaacgg ccgccagtgt gctggaattc gcccttgggg tctctgtcac gatacgcgta
      361 tcgatgcatc tacttttctc tatactgtac gtttcaatct ggggaagcgg aatcccaaaa
      421 gggaaagccg ccgcattaag ctccacagcg ggaagcggaa tcccaaaagg gaaagccgcc
      481 gcattaagct ccacagcggg aagcggaatc ccaaaaggga aagccgccgc attaagctcc
      541 acagcgggaa gcggaatccc aaaagggaaa gccgccgcat taagctccac agccttgcat
      601 aatccgatga cctgactagt gcggacaaag actattattt cgaggcaagg ccaccacgta
      661 ccgcggtccc aaacttttgc aaagctgaaa acagcgtggg ggtcaacgtg gatcagaaag
      721 aggggcagat cagcttctat aagaagctcc tttccccaca attggcccac acgacacttc
      781 tacacactta cacatctact ggatccccac agagaccgga agggcgaatt ctgcagatat
      841 ccatcacact ggcggccgct cgagcatgca tctagagggc ccaattcgcc ctatagtgag
      901 tcgtattaca attcactggc cgtcgtttta caacgtcgtg actgggaaaa ccctggcgtt
      961 acccaactta atcgccttgc agcacatccc cctttcgcca gctggcgtaa tagcgaagag
     1021 gcccgcaccg atcgcccttc ccaacagttg cgcagcctat acgtacggca gtttaaggtt
     1081 tacacctata aaagagagag ccgttatcgt ctgtttgtgg atgtacagag tgatattatt
     1141 gacacgccgg ggcgacggat ggtgatcccc ctggccagtg cacgtctgct gtcagataaa
     1201 gtctcccgtg aactttaccc ggtggtgcat atcggggatg aaagctggcg catgatgacc
     1261 accgatatgg ccagtgtgcc ggtctccgtt atcggggaag aagtggctga tctcagccac
     1321 cgcgaaaatg acatcaaaaa cgccattaac ctgatgttct ggggaatata aatgtcaggc
     1381 atgagattat caaaaaggat cttcacctag atccttttca cgtagaaagc cagtccgcag
     1441 aaacggtgct gaccccggat gaatgtcagc tactgggcta tctggacaag ggaaaacgca
     1501 agcgcaaaga gaaagcaggt agcttgcagt gggcttacat ggcgatagct agactgggcg
     1561 gttttatgga cagcaagcga accggaattg ccagctgggg cgccctctgg taaggttggg
     1621 aagccctgca aagtaaactg gatggctttc ttgccgccaa ggatctgatg gcgcagggga
     1681 tcaagctctg atcaagagac aggatgagga tcgtttcgca tgattgaaca agatggattg
     1741 cacgcaggtt ctccggccgc ttgggtggag aggctattcg gctatgactg ggcacaacag
     1801 acaatcggct gctctgatgc cgccgtgttc cggctgtcag cgcaggggcg cccggttctt
     1861 tttgtcaaga ccgacctgtc cggtgccctg aatgaactgc aagacgaggc agcgcggcta
     1921 tcgtggctgg ccacgacggg cgttccttgc gcagctgtgc tcgacgttgt cactgaagcg
     1981 ggaagggact ggctgctatt gggcgaagtg ccggggcagg atctcctgtc atctcacctt
     2041 gctcctgccg agaaagtatc catcatggct gatgcaatgc ggcggctgca tacgcttgat
     2101 ccggctacct gcccattcga ccaccaagcg aaacatcgca tcgagcgagc acgtactcgg
     2161 atggaagccg gtcttgtcga tcaggatgat ctggacgaag agcatcaggg gctcgcgcca
     2221 gccgaactgt tcgccaggct caaggcgagc atgcccgacg gcgaggatct cgtcgtgacc
     2281 catggcgatg cctgcttgcc gaatatcatg gtggaaaatg gccgcttttc tggattcatc
     2341 gactgtggcc ggctgggtgt ggcggaccgc tatcaggaca tagcgttggc tacccgtgat
     2401 attgctgaag agcttggcgg cgaatgggct gaccgcttcc tcgtgcttta cggtatcgcc
     2461 gctcccgatt cgcagcgcat cgccttctat cgccttcttg acgagttctt ctgaattatt
     2521 aacgcttaca atttcctgat gcggtatttt ctccttacgc atctgtgcgg tatttcacac
     2581 cgcatcaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta
     2641 aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataata
     2701 gcacgtgagg agggccacca tggccaagtt gaccagtgcc gttccggtgc tcaccgcgcg
     2761 cgacgtcgcc ggagcggtcg agttctggac cgaccggctc gggttctccc gggacttcgt
     2821 ggaggacgac ttcgccggtg tggtccggga cgacgtgacc ctgttcatca gcgcggtcca
     2881 ggaccaggtg gtgccggaca acaccctggc ctgggtgtgg gtgcgcggcc tggacgagct
     2941 gtacgccgag tggtcggagg tcgtgtccac gaacttccgg gacgcctccg ggccggccat
     3001 gaccgagatc ggcgagcagc cgtgggggcg ggagttcgcc ctgcgcgacc cggccggcaa
     3061 ctgcgtgcac ttcgtggccg aggagcagga ctgacacgtg ctaaaacttc atttttaatt
     3121 taaaaggatc taggtgaaga tcctttttga taatctcatg accaaaatcc cttaacgtga
     3181 gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc
     3241 tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt
     3301 ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc
     3361 gcagatacca aatactgttc ttctagtgta gccgtagtta ggccaccact tcaagaactc
     3421 tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg
     3481 cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg
     3541 gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga
     3601 actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc
     3661 ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg
     3721 gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg
     3781 atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt
     3841 tttacggttc ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc
     3901 tgattctgtg gataaccgta ttaccgcctt tgagtgagct gataccgctc gccgcagccg
     3961 aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa g
//
